Immunization with Plant-Expressed Hemagglutinin Protects Chickens from Lethal Highly Pathogenic Avian Influenza Virus H5N1 Challenge Infection

Abstract

Highly pathogenic avian influenza (HPAI) is a striking disease in susceptible poultry, which leads to severe economic losses. Inactivated vaccines are the most widely used vaccines in avian influenza virus (AIV) vaccination programs. However, these vaccines interfere with the serological detection of wild-type AIV infections in immunized populations. The use of vaccines that allow differentiation between infected and vaccinated animals (DIVA strategy) would stop current stamping-out policies. Therefore, novel vaccination strategies are needed to allow improved protection of animals and humans against HPAI virus (HPAIV) infection. The presented study analyzed for the first time the immunogenic capacity of plant-expressed full-length hemagglutinin (rHA0) of HPAIV H5N1 in several vaccine formulations within the highly relevant host species chicken. We were able to express plant-expressed rHA0 at high levels and could show that, when administered with potent adjuvants, it is highly immunogenic and can fully protect chicken against lethal challenge infection. Real-time reverse transcription (RT)-PCR and serological tests demonstrated only marginally increased virus replication in animals vaccinated with plant-derived rHA0 compared to animals immunized with an inactivated reference vaccine. In addition, the use of plant-expressed rHA0 also allowed an easy serological differentiation of vaccinated from AIV-infected animals based on antibodies against the influenza virus NP protein.Highly pathogenic avian influenza (AI) (HPAI) is a striking disease in susceptible poultry, which leads to severe economic losses (21). Since 2003, the H5N1 HPAI epidemic has claimed over 220 million poultry and other birds either through direct mortality from infection or from preemptive culling (22). The implementation of vaccination of poultry as a tool for the reduction of the viral load in the environment and, thus, for decreasing the risk of transmission within poultry-and, as a consequence, to humans-is still a discussed topic. Inactivated vaccines are the most widely used vaccines in AI vaccination programs. They are particularly addressed to protect adult chickens, turkeys, and other birds in emergency situations, e.g., when ring vaccination is used in an area of an HPAI virus (HPAIV) outbreak or when prophylactic vaccination is used in a region where H5 or H7 AI virus (AIV) infections are endemic. However, these vaccines limited the serological detection of wild-type AIV infections in immunized populations, as wild-type infection could be detectable only through higher antibody titers to nonstructural proteins or if the neuraminidase subtype of the vaccine differed from the subtype of the introduced wild-type virus (28).The use of vaccines that fit in any case to the strategy of differentiating infected from vaccinated animals (DIVA) would make a strong case for turning away from current stamping-out policies in many countries. Vaccines that consist of only one major antigenic protein related to influenza A virus (e.g., hemagglutini...

Transient Expression of an Immunogenic Envelope Attachment Glycoprotein of Nipah Virus in Nicotiana benthamiana

“…Other instances of successful expression of plant-based vaccine antigens or virus-like particles include foot and mouth disease virus structural protein VP1 expressed in transgenic alfalfa plants (Wigdorovitz et al 1999), chimeric peptide containing antigenic determinants from rabies virus glycoprotein and nucleoprotein expressed in Nicothiana sp. plants (Yusibov et al 2002), cholera toxin B subunit expressed in transgenic tobacco chloroplasts (Daniell et al 2001), recombinant subunit hemagglutinin of influenza A virus expressed in Nicothiana benthamiena (Kalthoff et al 2010;Mett et al 2008;Shoji et al 2011Shoji et al , 2008 and extracellular domain of matrix protein 2 (M2e) of influenza A virus expressed in Nicothiana benthamiena (Mardanova 2015).…”

Biotechnological leap in biopharmaceuticals: Current status and challenges ahead

“…The expression of recombinant hemagglutinin was demonstrated by Kalthoff et al [80]. This study was done by using N. benthamiana because N. benthamiana optimized expression levels [80]. This can be used for the prevention of chicken from very dangerous and life taking infections of heterologous highly pathogenic influenza viruses H5N1.…”

“…In the solution form the production of influenza virus can be quick and in a huge amount. The expression of recombinant hemagglutinin was demonstrated by Kalthoff et al [80]. This study was done by using N. benthamiana because N. benthamiana optimized expression levels [80].…”

The Role of the Plant-Specific ALTERED XYLOGLUCAN9 Protein in Arabidopsis Cell Wall PolysaccharideO-Acetylation

“…For the protease protection assays, the coding sequence of interest was cloned into pICH31070 (Icon Genetics) along with the appropriate accessory constructs (a plasmid with p19 for the fluorescence microscopy [Voinnet et al, 2003] and plasmids pICH20111 [Kalthoff et al, 2010] and pICH14011 [Giritch et al, 2006] for the protease protection assays). The plasmids containing the desired coding sequences were transformed into Agrobacterium tumefaciens strain GV3101 by electroporation.…”